Development of a novel and specialized cultivation method for isolating Helicobacter pullorum from chicken meat

Document Type : Short paper


1 DVM Student, Faculty of Veterinary Medicine, Semnan University, Semnan, Iran

2 Department of Clinical Science, Faculty of Veterinary Medicine, Semnan University, Semnan, Iran

3 Department of Food Hygiene and Quality Control, Faculty of Veterinary Medicine, Semnan University, Semnan, Iran


Background: It has become established that Helicobacter pullorum could be isolated from raw chicken meat. Aims: This study was aimed to develop a novel culture method (protocol B) to isolate H. pullorum from chicken meat by adding some modifications to the traditional culture method (protocol A), and as a consequence to compare their sensitivity, specificity, and the accuracy of these methods with polymerase chain reaction (PCR) test. Methods: 400 chicken meat samples were collected from various retail markets and supermarkets. Each sample was processed by protocol A, protocol B, and PCR test. Results: Out of 400 samples, 77 (19.25%), and 163 (40.75%) were culture-positive by protocol A and protocol B, respectively. Using PCR test as a gold standard, 196 (49%) samples were identified as H. pullorum. The specificity for both protocols was determined 100%, while the sensitivity of protocol B and protocol A was assessed 83% and 39%, respectively. Also, the higher and lower accuracy belonged to protocol B (92%) and protocol A (70%), respectively. Conclusion: The methodology designed herein can provide a suitable, approximately sensitive, specific, and accurate method to cultivate H. pullorum from chicken meat.


Main Subjects

Amjad, M (2020). An overview of the molecular methods in the diagnosis of gastrointestinal infectious diseases. Int. J. Microbiol., 2020: 1-13.
Behroo, S; Javadi, A and Rad, MG (2015). Helicobacter pullorum prevalence in patients with gastroenteritis in humans and chicken in the province of Ardabil in 2014. Indian J. Fundam. Appl. Life Sci., 5: 87-94.
Bello, N; Francino, O and Sánchez, A (2001). Isolation of genomic DNA from feathers. J. Vet. Diagnostic. Investig., 13: 162-164.
Bojanić, K; Midwinter, AC; Marshall, JC; Biggs, PJ and Acke, E (2019). Isolation of emerging Campylobacter species in working farm dogs and their frozen home-killed raw meat diets. J. Vet. Diagnostic. Investig., 31: 23-32.
Borges, V; Santos, A; Correia, CB; Saraiva, M; Menard, M; Vieira, L; Sampaio, DA; Pinheiro, M; Gomez, GB and Olestro, M (2015). Helicobacter pullorum isolated from fresh chicken meat: Antibiotic resistance and genomic traits of an emerging foodborne pathogen. Appl. Environ. Microbiol., 81: 8155-8163.
Ceelen, L; Decostere, A; Verschraegen, G; Ducatelle, R and Haesebrouck, F (2005). Prevalence of Helicobacter pullorum among patients with gastrointestinal disease and clinically healthy persons. J. Clin. Microbiol., 43: 2984-2986.
Ceelen, MA and Haesebrouck, F (2005). In vitro susceptibility of Helicobacter pullorum strains to different antimicrobial agents. Microb. Drug Resist., 11: 122-126.
Deshmukh, D; Joseph, J; Chakrabarti, M; Sharma, S; Jayasudha, R; Sama, KC; Sontam, B; Tyagi, M; Narayana, R and Shivaji, S (2019). New insights into culture negative endophthalmitis by unbiased next generation sequencing. Sci. Rep., 9: 844-853.
El-shibiny, A; Connerton, P and Connerton, I (2009). Survival at refrigeration and freezing temperatures of Campylobacter coli and Campylobacter jejuni on chicken skin applied as axenic and mixed inoculums. Int. J. Food. Microbiol., 131: 197-202.
González, A; Piqueres, P; Moreno, Y; Canigral, I; Owen, RG; Hernandez, J and Ferrus, MA (2008). A novel real-time PCR assay for the detection of Helicobacter pullorum-like organisms in chicken products. Int. Microbiol., 11: 203-208.
Hassan, AK; Shahata, MA; Rafaie, EM and Ibrahim, RS (2014). Pathogenesity testing and antimicrobial susceptibility of H. pullorum isolates from chicken origin. Int. J. Vet. Sci. Med., 2: 72-77.
Jamshidi, A; Bassami, RM; Salami, H and Mohammadi, S (2014). Isolation and identification of Helicobacter pullorum from caecal content of broiler chickens in Mashhad, Iran. Iran. J. Vet. Res., 15: 179-182.
Javed, K; Gul, F; Abbasi, R; Zaidi, RA; Noreen, Z; Bokhari, H and Javed, S (2019). Prevalence and role of type six secretion system in pathogenesis of emerging zoonotic pathogen Helicobacter pullorum from retail poultry. Avian Pathol., 48: 557-563.
Javed, S; Gul, F; Javed, K and Bokhari, H (2017). Helicobacter pullorum: An emerging zoonotic pathogen. Front. Microbiol., 8: 604-611.
Jebellijavan, A; Emadi Chashmi, SH; Staji, H and Akhlaghi, H (2020). Comparison of the culture and PCR methods to determine the prevalence and antibiotic resistance of H. pullorum from chicken thigh samples in Semnan, Iran. J. Hum. Environ. Health Promot., 6: 167-172.
Kahraman, BB and Seyyal, AK (2013). Investigation of Helicobacter pullorum occurence in chicken in the marmara region of Turkey. Istanbul. Univ. Vet. Fak. Derg., 39: 63-66.
Kim, J; Shin, H; Park, H; Shin, H; Park, H; Jung, H; Kim, J; Cho, S; Ryu, S and Jeon, B (2019). Microbiota analysis for the optimization of Campylobacter isolation from chicken carcasses using selective media. Front. Microbiol., 10: 1381-1389.
Manfreda, G; Parisi, A; Lucchi, A; Zanoni, RG and De cesare, A (2011). Prevalence of Helicobacter pullorum in conventional, organic, and free-range broilers and typing of isolates. Appl. Environ. Microbiol., 77: 479-484.
Manfreda, G; Rossi, M; Sanguinetti, V; Gavioli, R; Lozito, P; De cesare, A and Zanoni, RG (2006). Prevalence of Helicobacter pullorum in broiler chickens reared in intensive and extensive farms. Appl. Environ. Microbiol., 77: 479-484.
Mohamed, M; Ragab, I; Shahata, M and El-refaie, M (2010). Helicobacter pullorum among poultry in ASSIUT-Egypt: Genetic characterization, virolence and MIC. Int. J. Poult. Sci., 9: 521-526.
Molla Kazemiha, V; Amanzadeh, A; Memarnejadian, A; Azari, S; Shokrgozar, MA; Mahdian, R and Bonakdar, S (2014). Sensitivity of biochemical test in comparison with other methods for the detection of mycoplasma contamination in human and animal cell lines stored in the National Cell Bank of Iran. Cytotechnology. 66: 861-873.
Ochoa, S; Martínez, OA; Fernández, H and Collado, L (2019). Comparison of media and growth conditions for culturing enterohepatic Helicobacter species. Lett. Appl. Microbiol., 69: 190-197.
Qumar, S; Majid, M; Kumar, N; Tiwari, SK; Semmler, T; Devi, S; Baddam, R; Hussain, A; Shaik, S and Ahmed, N (2017). Crossm genome dynamics and molecular resistant Helicobacter pullorum isolates obtained from broiler and free-range chickens in India. Appl. Environ. Microbiol., 83: e02305.
Stanley, J; Linton, D; Burnes, AP; Dehwirst, FE; On, SLW; Porter, A; Owen, RG and Costas, M (1994). Helicobacter pullorum sp. nov. genotype and phenotype of
a new species isolated from poultry and from human patients with gastroenteritis. Microbiology. 140: 3441-3449.
Tilmanne, A; Marisca, H; Yattara, K; Herpol, M; Vlaes, L; Retore, P; Quach, C; Vandenberg, O; Hallin, M and Martiny, D (2019). Multi-step optimization of the filtration method for the isolation of Campylobacter species from stool samples. Eur. J. Clin. Microbiol. Infect. Dis., 38: 859-864.
Wafaa, A and El-ghany, A (2020). Helicobacter pullorum: A potential hurdle emerging pathogen for public health. J. Inf. Develop. Countries. 14: 1225-1230.
Wai, SS; Abdul-aziz, S; Bitrus, AA; Zunita, Z and Abu, J (2019). Helicobacter pullorum in broiler chickens and the farm environment: A one health approach. Int. J. One Heal. 5: 20-25.
Zanoni, RG; Piva, S; Rossi, M; Pasquali, F; Lucci, A; De cesare, A and Manfreda, G (2011). Occurrence of Helicobacter pullorum in turkeys. Vet. Microbiol., 149: 492-496.
Zanoni, RG; Rossi, M; Giacomucci, D; Sanguinetti, V and Manfreda, G (2007). Occurrence and antibiotic susceptibility of Helicobacter pullorum from broiler chickens and commercial laying hens in Italy. Int. J. Food. Microbiol., 116: 168-173.